The insulin receptor is encoded by a single copy gene located on chromosome 19 in the human. In addition, at least two other highly homologous genes have been identified. One gene encodes the receptor for insulin like growth factors (IGF)-I and -II. The third gene in the family encodes another receptor tyrosine kinase. The ligand for this third receptor has not been identified; however, preliminary evidence suggests that this insulin-receptor related receptor (IRR) does not bind insulin, IGF-I, or IGF-II. The goals of this project are two-fold: (1) to identify the ligand for this receptor, and (2) to elucidate the physiological role of the IRR. Toward this end, we have cloned human IRR cDNA. This work revealed that the cDNA undergoes differential splicing; some of the transcripts (<10%) use an alternate splice acceptor site that is located 24 nucleotides upstream from the predicted 5'-end of exon 14. Use of this alternative splice acceptor site preserves the reading frame in human IRR cDNA, this would not be the case in the mouse gene since the site is located 23 nucleotides upstream from exon 14. In addition, multiple alternative splicing patterns were identified in exons 1 and 2 of the murine gene. None of these preserve the reading frame. The function of these alternative splicing patterns has not been elucidated. By expressing IRR cDNA in cultured cells, we hope to develop a bioassay for the IRR ligand. In addition, we have cloned the murine IRR gene. A fragment of the cloned gene has been used to construct a targeting vector that is being used to inactivate the IRR gene by homologous recombination in embryonic stem cells. Once this is successful, we will attempt to construct transgenic mice with mutations in the IRR gene. These transgenic mice will be studied in an effort to determine the phenotypic effect of mutations in the IRR gene. This has the potential to provide clues into the physiological roles of the IRR and its ligand.